Spinorphin (LVVYPWT) has been isolated from the bovine spinal cord as an endogenous inhibitor of enkephalin-degrading enzymes. It has been reported that spinorphin has an antinociceptive effect, inhibitory effect on contraction of smooth muscle and anti-inflammatory effect. In the present study, the effects of leu-enkephalin and spinorphin on allodynia and mechanical and thermal nociceptions were examined in vivo using mice. Intrathecal (i.t.) administration of leu-enkephalin or spinorphin inhibited the allodynia induced by intrathecal nociceptin in a dose-dependent manner. Furthermore, spinorphin enhanced the inhibitory effect of enkephalin on allodynia induced by nociceptin. Naloxone antagonized both inhibitory effects of leu-enkephalin and spinorphin, suggesting that the endogenous opioidergic system can modulate allodynia. Intracerebroventricular (i.c.v.) administration of leu-enkephalin increased the nociceptive threshold of heat or mechanical stimulation to a mouse. Although i.c.v. administration of spinorphin had no effect on the threshold of heat or mechanical stimulation, spinorphin enhanced and prolonged the antinociceptive effect of leu-enkephalin. The enhancement of spinorphin on the antinociception produced by leu-enkephalin was reversed by pretreatment with naloxone. From these results, it is suggested that the effects of spinorphin on enkephalin-induced anti-allodynic and antinociceptive effects are due to inhibition of enkephalin-degrading enzymes.